Multiwell plate devices serve a broad spectrum of laboratory uses. Most applications involve attachment or immobilization of biological materials including, without limitation, biomolecules such as polypeptides and nucleic acids, cells, tissues or fragments biological material, to a surface within the wells (sidewall and/or bottom surface) and the performance of one or more reactions followed by some sort of quantitative and/or qualitative analytical process.
Robotic instruments have been developed for performing automated processing of multiwell plates. Such automated processes include, without limitation, deposition of biological materials (spotting, printing, etc.), addition or removal of reagents, washing, scanning and analysis. The capability of such automated instruments is typically limited to processing plates with “standard” dimensions as established by the Society of Biomolecular Sciences (SBS Standards). Thus, the “footprint” for most multiwell plates is approximately 85 mm×125 mm with wells located in a standardized format depending upon the total number of wells. The American National Standards Institute (ANSI) has published the SBS Standards for microplates as: “Footprint Dimensions” (ANSI/SBS 1-2004), “Height Dimensions” (ANSI/SBS 2-2004), “Bottom Outside Flange Dimensions” (ANSI/SBS 3-2004) and “Well Postions” (ANSI/SBS 4-2004). All of these ANSI/SBS publications are incorporated herein by reference.
Although a standard structure for multiwell plates has facilitated automatic robotic processing, at the same time the structure presents a challenge with regards to certain types of procedures, particularly as the number of wells grows beyond 96. For example, spotting or printing of a microarray on the bottom surface of a well using automatic/robotic liquid handling systems or “arrayers” requires the pin or stylus or other printing/spotting means to move significantly up and down as it arrays one well and moves to the next to print or spot another array. Such movement increases processing time and increases the risk of damage to printing pins or stylus from unwanted collision with plate features above the surface to be printed or arrayed. Therefore, a need exists for a multiwell plate device more conducive to rapid processing.
Moreover, analysis of reactions occurring in the wells of a multiwell plate presents a challenge. Often, the analysis is accomplished by detecting or measuring a change in the material attached to the bottom surface of the wells (substrate) rather than a change in a fluid reaction mixture contained within the wells, as is the case for ELISA-type assays.
Optical detection is the most commonly utilized method to detect changes in surface-localized reactions, particularly with regards to arrays representing multiple different reactions. For surface-localized reactions, the focal plane for proper measurement of the reaction is often limited to a very small range of depths, typically a range of no more than about 5 mm. Analysis, whether done via automated scanning or microscopy or other means, can be performed by directing a light or energy source from above the reaction surface of the substrate or from below (through the substrate) and focusing an optic that captures the detectable signal from above or below the reaction surface. In some cases, for example when certain types of coated substrates and/or mixtures of detection agents are used, analysis from both above and below the reaction surface is useful in order to glean optimal data. However, the design of a standard multi-well plate complicates efforts to analyze results from both above and below the reaction surface. Most automatic scanners/analyzers can scan from only above or below the reaction surface but not both. Because of the dimensions of a standard multiwell plate, the focal plane of the reaction surface (bottom surface of the wells) when the plate is upright is significantly different from the focal plane when the plate is turned over. One prior art solution has been to use two separate analysis systems wherein one is capable of scanning from above the reaction surface and the other from below. Such an approach is expensive and time-consuming. Alternatively, another solution has been use of a multiwell plate device comprising separate pieces assembled to form the plate including a substrate that is detachable from the multiwell plate structure to eliminate physical interference by the plate structure with the focal plane of the reaction surface.
U.S. patent application Ser. No. 10/739,784 to Harvey et al., incorporated herein by reference, teaches the use of 1-4 glass microscope slides placed into a frame-like holder having standard multiwell plate dimensions. The slides are spotted or printed prior to placement in the holder. Once in place in the holder, each slide is topped in a releasable manner with a separate multiwell chamber plate having bottomless wells such that the printed glass slide forms a bottom surface for the chamber plate. Finally a retention means is used to retain the slides in the holder. After processing, the chamber plates and slides are removed from the holder and separated, and each slide is analyzed. Thus, the frame-like holder is used only during the reaction phase of the process; the steps of printing arrays and analyzing results are performed on each individual slide while separated from the holder.
U.S. patent application Ser. No. 11/134,449 to Haines et al., incorporated herein by reference, teaches a device comprising a substrate with a functional coating and biomolecules attached thereto. The substrate is reversibly attached to a superstructure containing multiple openings (multiwell structure). A frame-like tray holds the substrate and serves as an alignment jig for the superstructure. After processing, the system is completely disassembled to remove the substrate for analysis. Thus, the assembled device is used during the reaction phase of the procedure and, optionally, during the step of printing arrays, but it is disassembled for analysis.
U.S. Pat. No. 7,063,979 to MacBeth et al., incorporated herein by reference, teaches a microtiter-microarray device comprising a bottomless multiwell plate structure, one or more substrates having predeposited microarrays, and one or more gaskets for sealing the substrates to the multi-well plate structure. The seal must be fluid-tight but may be either reversible or irreversible. The patent teaches use of a first aligning device to align the gasket and plate structure for attachment purposes and a second aligning device for attachment of the substrates bearing predeposited microarrays. A separate device is used to remove the substrate after processing for analysis via conventional slide scanner. Alternatively, the substrate can remain attached to the gasket and plate structure for analysis via plate scanner, for example, Tecan LS-200 scanner (Tecan, Durham, N.C.). Thus, the reaction surface in a fully assembled multiwell plate device falls within a particular focal plane when the plate is upright and a significantly different focal plane when turned over. As described in U.S. Pat. No. 7,063,979, to scan the reaction surface from the opposite side with a plate scanner, the substrate must be detached and turned over 180°.
A detachable substrate presents a challenge because it must be attached to the plate structure in such a way as to be fluid-tight during the reaction phase of processing and yet removable without a level of force that could break or otherwise damage the substrate and without leaving adhesive or other material that might interfere with analysis. A need exists for a multiwell plate device wherein the reaction surface can be scanned from above or below while maintained within the detectable focal plane of a scanning device without requiring detachment of the substrate from the multiwell plate structure.